title: Stimulated emission depletion-based raster image correlation spectroscopy reveals biomolecular dynamics in live cells.
abstract: Raster image correlation spectroscopy is a powerful tool to study fast molecular dynamics such as protein diffusion or receptor-ligand interactions inside living cells and tissues. By analysing spatio-temporal correlations of fluorescence intensity fluctuations from raster-scanned microscopy images, molecular motions can be revealed in a spatially resolved manner. Because of the diffraction-limited optical resolution, however, conventional raster image correlation spectroscopy can only distinguish larger regions of interest and requires low fluorophore concentrations in the nanomolar range. Here, to overcome these limitations, we combine raster image correlation spectroscopy with stimulated emission depletion microscopy. With imaging experiments on model membranes and live cells, we show that stimulated emission depletion-raster image correlation spectroscopy offers an enhanced multiplexing capability because of the enhanced spatial resolution as well as access to 10-100 times higher fluorophore concentrations.
authors.affiliation: Institute of Applied Physics and Center for Functional Nanostructures, Karlsruhe Institute of Technology, 76128 Karlsruhe, Germany.
authors.forename: Per Niklas
authors.initials: PN
authors.lastname: Hedde
authors.forename: René M
authors.initials: RM
authors.lastname: Dörlich
authors.forename: Rosmarie
authors.initials: R
authors.lastname: Blomley
authors.forename: Dietmar
authors.initials: D
authors.lastname: Gradl
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Oppong
authors.forename: Andrew C B
authors.initials: AC
authors.lastname: Cato
authors.forename: G Ulrich
authors.initials: GU
authors.lastname: Nienhaus
mesh_terms: D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D004058:Diffusion; D008051:Lipid Bilayers / Q000737:chemistry; D050356:Lipid Metabolism; D046911:Macromolecular Substances / Q000378:metabolism*; D018613:Microscopy, Confocal / Q000379:methods*; D008954:Models, Biological; D010714:Phosphatidylethanolamines / Q000737:chemistry; D051381:Rats; D013050:Spectrometry, Fluorescence / Q000379:methods*; D014981:Xenopus
chemical_list: D008051:Lipid Bilayers; D046911:Macromolecular Substances; D010714:Phosphatidylethanolamines; C007510:1,2-dipalmitoyl-3-phosphatidylethanolamine